Search

We have recently identified a pool of intracellular β1 adrenergic receptors (β1ARs) at the sarcoplasmic reticulum (SR) crucial for cardiac function. Here, we aim to characterize the integrative control of intracellular catecholamine for subcellular β1AR signaling and cardiac function. Using anchored Förster resonance energy transfer (FRET) biosensors and transgenic mice, we determined the regulation of compartmentalized β1AR-PKA signaling at the SR and plasma membrane (PM) microdomains by organic cation transporter 3 (OCT3) and monoamine oxidase A (MAO-A), two critical modulators of catecholamine uptake and homeostasis. Additionally, we examined local PKA substrate phosphorylation and excitation-contraction coupling in cardiomyocyte. Cardiac-specific deletion of MAO-A (MAO-A-CKO) elevates catecholamines and cAMP levels in the myocardium, baseline cardiac function, and adrenergic responses. Both MAO-A deletion and inhibitor (MAOi) selectively enhance the local β1AR-PKA activity at the SR but not PM, and augment phosphorylation of phospholamban, Ca2+ cycling, and myocyte contractile response. Overexpression of MAO-A suppresses the SR-β1AR-PKA activity and PKA phosphorylation. However, deletion or inhibition of OCT3 by corticosterone prevents the effects induced by MAOi and MAO-A deletion in cardiomyocytes. Deletion or inhibition of OCT3 also negates the effects of MAOi and MAO-A deficiency in cardiac function and adrenergic responses in vivo. Our data show that MAO-A and OCT3 act in concert to fine-tune the intracellular SR-β1AR-PKA signaling and cardiac fight-or-flight response. We reveal a drug contraindication between anti-inflammatory corticosterone and anti-depressant MAOi in modulating adrenergic regulation in the heart, providing novel perspectives of these drugs with cardiac implications.

Spatiotemporal regulation of subcellular protein kinase A (PKA) activity for precise substrate phosphorylation is essential for cellular responses to hormonal stimulation. Ryanodine receptor 2 (RyR2) and (sarco)endoplasmic reticulum calcium ATPase 2a (SERCA2a) represent two critical targets of β adrenoceptor (βAR) signaling on the sarcoplasmic reticulum membrane for cardiac excitation and contraction coupling. Using novel biosensors, we show that cardiac β1AR signals to both RyR2 and SERCA2a nanodomains in cardiomyocytes from mice, rats, and rabbits, whereas the β2AR signaling is restricted from these nanodomains. Phosphodiesterase 4 (PDE4) and PDE3 control the baseline PKA activity and prevent β2AR signaling from reaching the RyR2 and SERCA2a nanodomains. Moreover, blocking inhibitory G protein allows β2AR signaling to the RyR2 but not the SERCA2a nanodomains. This study provides evidence for the differential roles of inhibitory G protein and PDEs in controlling the adrenergic subtype signaling at the RyR2 and SERCA2a nanodomains in cardiomyocytes. 6RLYcPB_fJ7pnT18c_tdPo Video abstract Highlights: Design a FRET-based biosensor to monitor PKA dynamics at the RyR2 nanodomains in myocytes Stimulation of β1AR promotes PKA activity at both RyR2 and SERCA2a nanodomains whereas stimulation of β2AR does not Inhibition of PDE3 and PDE4 enhances PKA activity at the baseline and after β2AR stimulation Inhibition of Gi selectively permits β2AR signaling to the RyR2 nanodomains but not SERCA2a nanodomains. [ABSTRACT FROM AUTHOR]

We have recently identified a pool of intracellular β1 adrenergic receptors (β1ARs) at the sarcoplasmic reticulum (SR) crucial for cardiac function. Here, we aim to characterize the integrative control of intracellular catecholamine for subcellular β1AR signaling and cardiac function. Using anchored Förster resonance energy transfer (FRET) biosensors and transgenic mice, we determined the regulation of compartmentalized β1AR-PKA signaling at the SR and plasma membrane (PM) microdomains by organic cation transporter 3 (OCT3) and monoamine oxidase A (MAO-A), two critical modulators of catecholamine uptake and homeostasis. Additionally, we examined local PKA substrate phosphorylation and excitation–contraction coupling in cardiomyocyte. Cardiac-specific deletion of MAO-A (MAO-A-CKO) elevates catecholamines and cAMP levels in the myocardium, baseline cardiac function, and adrenergic responses. Both MAO-A deletion and inhibitor (MAOi) selectively enhance the local β1AR-PKA activity at the SR but not PM, and augment phosphorylation of phospholamban, Ca2+ cycling, and myocyte contractile response. Overexpression of MAO-A suppresses the SR-β1AR-PKA activity and PKA phosphorylation. However, deletion or inhibition of OCT3 by corticosterone prevents the effects induced by MAOi and MAO-A deletion in cardiomyocytes. Deletion or inhibition of OCT3 also negates the effects of MAOi and MAO-A deficiency in cardiac function and adrenergic responses in vivo. Our data show that MAO-A and OCT3 act in concert to fine-tune the intracellular SR-β1AR-PKA signaling and cardiac fight-or-flight response. We reveal a drug contraindication between anti-inflammatory corticosterone and anti-depressant MAOi in modulating adrenergic regulation in the heart, providing novel perspectives of these drugs with cardiac implications. [ABSTRACT FROM AUTHOR]